A number of plaque morphology (syn) mutants of herpes simplex virus type 1 (HSV-1) have been isolated which, unlike wild type virus, cause the formation of extensive syncytia in an otherwise normal infection. Although synctia are not formed in wild type infections a limited amount of fusion is observed. Fusion starts in wild type infected cells but stops a short time later, perhaps because of the appearance of putative inhibitor glycoproteins on the cell surface. The syn mutants isolated to date may have altered inhibitor glycoproteins because the appearance of these glycoproteins on the cell surface is delayed. To genetically identify fusion factor and inhibitor genes more syn mutants and revertants of syn mutants will be isolated. Revertants will be analyzed for the presence of a second mutation which could be in the fusion factor gene. These revertants would contain suppressor mutations of cell fusion. Since fusion activity may be required for virus penetration of the host cell it is also proposed to isolate ts penetration mutants from syn mutants and test their ability to cause cell fusion at the nonpermissive temperature. To biochemically identify possible fusion factors and inhibitors during infection with wild type and syn mutant viruses plasma membranes, isolated nuclei and whole cell extracts will be examined using two-dimensional gel electrophoresis for differences in the time of appearance and amounts of protein and glycoproteins. In addition, the proteins and glycoproteins of the corresponding virions will be examined. It is proposed to develop an in vitro fusion assay for use as a model system to study the molecules that affect cell fusion. We plan to isolate the putative inhibitor glycoproteins and characterize their peptide fingerprints and oligosaccharide chain lengths and composition. It is also proposed to test their effect on cell fusion in the in vitro fusion assay system.